Grant Number: 1DP2AI138259 - 01 PI Name: Malvankar, Nikhil Revised Project abstract: Chronic bacterial infections pose dangerous health risks because they often require rigorous treatment regimens or surgeries. Current anti-microbials have little effect against persistent infections, and a key challenge to drug development is that bacterial survival mechanisms are not well understood. Most drugs target intracellular processes important for bacterial viability, but pathogens rapidly adapt and develop antibiotic resistance. To overcome these limitations, we aim to target extracellular charge interactions important for bacterial virulence to reduce selection pressure for drug-resistant mutations. Geobacter sulfurreducens use hair-like filaments called pili as ?nanowires? to transfer electrons for respiration and biofilm formation. We will evaluate whether pili of pathogens, which are crucial for lung infections of Cystic Fibrosis (CF) patients, show high conductivity similar to G. sulfurreducens pili. We have developed new tools to directly image and measure electrical charges and electron transfer in pili and living biofilms. By extending these tools to simultaneously analyze the interactions between host surface and living pathogens, we aim to identify the mechanism of infection by investigating three very common bacterial survival strategies, which are independent of each other, but cannot be explained fully by existing models: (1) Adhesion to host cells prevents bacteria from being washed away by bodily fluids, and is one of the most common microbial survival strategies. Furthermore, bacteria can adhere to each other to form biofilms, which block the efficacy of most anti-microbials, cause 80% of microbial infections in the body, and result in chronic infection and the need for surgical removal of afflicted areas. Experiments have shown that bacteria use pili for adhesion, but the biophysical mechanism is not clear. By correlated imaging of adhesion force and charge, we will determine the role of charge interactions in bacterial adhesion to the host cell. (2) Iron accumulation and metabolism of pathogens, particularly accumulation of Fe (II) during infection, hinders existing chelation therapies that target chelation of Fe (III). We will evaluate how bacterial charge interactions affect their iron accumulation and metabolism in CF lung. (3) Adaptation by pathogens in dynamic environments is a major factor in the failure of antimicrobial therapies, but the mechanisms of adaptation remain unclear. We will determine how charge interactions lower pH, mimicking the environment in the CF lung. Using newly-developed tools, we aim to achieve a comprehensive understanding of charge interactions to bring about a major shift in our understanding of bacterial infections. By identifying common bacterial survival mechanisms, targeted drugs can be developed to suppress infections.